86 LIFESTYLE COSMETICS
amounts of rutin, chlorogenic acid and salidroside was confirmed by dereplication analysis (Fig 3B and not shown). HPTLC also demonstrated adequate heat and storage stability. Heating to 80°C for 2 h or storage at 40°C for 3 months had no effect on the overall flavonoid pattern (Fig 3C).
Increasing the number of mitochondria and improving energy supply Objective To show that the generation of mitochondria is stimulated.
Technique
Mitochondria Assay: human fibroblasts were incubated with test substances or not (untreated) for 72 hours, followed by staining with a mitochondria specific dye. The intracellular fluorescence was measured in multiple overlapping areas and normalised to protein content, which is a marker for the number of cells present in each culture. Increased fluorescence points to elevated numbers of mitochondria per cell.
Results
The active ingredient stimulates mitochondrial biogenesis: the number of mitochondria per cell increased by a significant ~20% (Fig 4B). Every cell of the skin is thus equipped with an extra portion of powerhouses. This, in turn, will increase the energy level of the skin. Indeed, 0.01% of the active ingredient significantly increased the ATP level per cell by a significant ~20% (data not shown).
Improving the efficiency of energy production Objective To show that mitochondrial ROS emission is reduced. Harmful oxygen species are formed during energy generation in mitochondria. Increased mitochondrial numbers with increased energy production could consequently exhibit more oxidative stress.
Technique
DCF assay: ROS release was investigated by incubating human fibroblasts with test substances or not (untreated) for 72 h, followed by the addition of a non- fluorescing preliminary state of a dye that diffuses into the cell and is oxidised through ROS into a fluorescing product. A lower fluorescence signal thus indicates reduced mitochondrial ROS.
Results
The active ingredient provided additional energy while minimising side effects: skin cells with an increased number of mitochondria and higher energy production (see above) did not suffer from exaggerated
PERSONAL CARE EUROPE 3 *** 2
4
3
2
1
0 0 10 20 Time (min)
n Untreated n 0.01% n Quercetin n 0.001% n 0.1% HerbaGlow NRG
4 ** -18% *** -25% *** -29% 30 40
1
0 Untreated Quercetin 0.001 0.01 HerbaGlow NRG (%) 0.1
Figure 5: Reducing collateral ROS emissions. Skin cells were left untreated or were incubated with active ingredient or 3 ug/ml Quercetin (positive control24
). ROS release was determined over time (top)
as well as at the end of the assay (bottom). N = 12 from two independent experiments; Mean + SEM; Student’s unpaired t-test versus untreated control; * = p < 0.05; ** = p < 0.01; *** = p < 0.001.
ROS formation. The active ingredient even reduced free radical emission significantly and in a dose-dependent manner by up to ~30% indicating that energy production runs more efficiently (Fig 5). A more effective generation of energy with lower ROS emission is expected to result in a more youthful skin with recaptured glow and radiance.
Improving skin glow and radiance in vivo Objective To show that the active ingredient revives
the skin and offers glow and vitality. Skin with more energy is thought to be more radiant, which is a mirror of youth, health and happiness. Fabulous luminosity, homogenous complexion without imperfections, and smooth texture are all hallmarks of skin radiance.25,26 There is no universal definition of skin radiance, but it appears to be the combination of: 1. Level of skin uniformity, including texture (smooth skin), colour and imperfections (no spots). Here, we show skin smoothness as evaluated by Primos.
November 2018
ROS Emission (Normalized DCF Fluorescence)
ROS Emission (Normalized DCF Fluorescence)
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